B. subtilis has a multigene APase family of at least 5 APase genes. An APase multigene family has previously been observed only in mammals. Biochemical and genetic studies of the Bacillus APase family have identified APase gene subfamilies based on induction by phosphate starvation or sporulation induction or both. The long-term goal of this proposal is to understand the physiological function of each member of the APase gene family. The fact that the various APases of B. subtilis are located in different places in the cell, are produced at different times during development, and have different specific activities and substrate specificities implies that each isozyme has a unique function in the cell. Bacillus provides an excellent model system, combining powerful genetics with a developmental system to assess the roles of phosphatases. A prototype APase for each APase subfamily is being analyzed to determine: 1) mechanism of regulation of expression, 2) the final destination of the mature protein, and 3) protein structure of the APase. The regulation studies will determine in which cell type (sporulating or vegetative) APase has a functional role. The localization data will indicate a role in the mother cell or forespore if the APase is expressed in the sporulating cell, or an extracellular or intracellular role if the APase is expressed in the vegetative phosphate- starved cell. Study of APase structure will allow design of experiments to determine whether the surface domains of the protein have roles in protein localization or protein-protein interactions, factors that could determine substrate specificity and physiological function. Another long-term goal is to characterize the components of the pho regulon in Bacillus. This information is necessary to understand the regulation of the APase gene family and determine if the pho regulon or Pi concentration has a role in sporulation.